Efficacy of Kami Guibi-tang as an Add-On Therapy to Acetylcholinesterase Inhibitor for Cognitive Function in Mild Alzheimer's Disease: A Pilot Study

Background Kami Guibi-tang (KGT), a traditional Korean herbal medicine is mainly used to treat insomnia and nervousness. Acetylcholinesterase inhibitors (AChEIs) are the main treatments for mild Alzheimer's disease (AD), a degenerative brain disease. However, currently no drug can fundamentally treat AD or reverse the advanced cognitive decline. This clinical study explored the efficacy and safety of adding KGT to AChEI for cognitive function in mild AD. Methods This was a pilot study for a larger randomized, double-blind, placebo-controlled trial. Participants between 55–90 years diagnosed with mild AD were recruited from Kyung Hee University Hospital at Gangdong, Seoul, Korea. They were randomized to receive either KGT or placebo for 24 weeks, in addition to their regular AChEI. The primary outcome was treatment efficacy, as assessed by the relative amount of change over the study period in total scores on the Dementia version of the Seoul Neuropsychological Screening Battery (SNSB-D). Changes in SNSB subscores were assessed as secondary outcomes. Safety parameters, including adverse events and abnormalities in blood tests, electrocardiograms, and brain magnetic resonance imaging were also monitored. Results Between March 2018 and November 2020, seven participants each in the KGT group and the placebo group completed the 24-week trial. There were no significant changes in SNSB-D total or subindex scores for either group (p = 0.69 and 0.63, respectively), and no significant differences were observed between them (p=0.71). No adverse events related to KGT were reported. We also compared and analyzed the results of a previous pilot study conducted on amnestic mild cognitive impairment (aMCI) using protocol of this study. The aMCI group showed a significant improvement in the total SNSB-D score, especially in the memory domain, compared to the mild AD group (p = 0.04 and 0.02, respectively). The Korean Mini-Mental State Exam and Korean Instrumental Activities of Daily Living scores also significantly improved in the aMCI group (p = 0.01 and 0.02, respectively). Conclusions Compared to placebo, adding KGT to AChEI did not significantly improve cognitive function in SNSB in patients with mild AD. We suggest that KGT would have a positive effect on patients with early stages of cognitive impairment such as aMCI. The findings could assist design larger, longer-term clinical trials of KGT use in elderly patients with mild AD. This study was registered in the Korean Clinical Trial Registry on December 26, 2017, with the CRIS approval number KCT0002904.


Introduction
Alzheimer's disease (AD) accounts for 60-80% of all cases of dementia [1]. AD is a chronic progressive neurodegenerative disease characterized by memory impairment, gradual deterioration of other cognitive functions, and eventual loss of the ability to perform the functions of daily life. Tus, it afects the well-being of individuals in very profound ways [2].
Te number of dementia patients among people aged 65 years and older in Korea has been estimated at 860,000 as of 2019, or approximately 11% of the population. Te total number of afected people is expected to exceed one million by 2024 and further to three million by 2050 [3]. Te annual economic loss due to dementia is estimated at over 800 billion dollars worldwide, which is larger than that of any other degenerative brain disease [4]. Dementia also has a serious impact on the mental health and quality of life of family members. Terefore, early diagnosis and treatment is important to enable patients and their families to reduce avoidable medical expenses and prepare for various obstacles caused by dementia [3].
Te pathogenesis of AD has not yet been fully elucidated. Aggregation of amyloid beta (Aβ) and tau protein in the brain have been proposed as pathological biomarkers for AD [5]. A recent study showed that severely reactive astrocytes contribute to the development of AD [6]. However, no drugs are currently available that can fundamentally treat AD or help patients recover from advanced cognitive decline. Available drugs are mainly used to slow the progression of the disease and maintain cognitive function to the extent possible. In Korea, drugs approved for AD include acetylcholinesterase inhibitors (AChEIs), such as donepezil, rivastigmine, and galantamine, and N-methyl-D-aspartate (NMDA) receptor antagonists, such as memantine. However, in some patients, these drugs cause gastrointestinal side efects, such as nausea, diarrhea, and vomiting [7]. To overcome the limitations of the existing treatments, studies with various complementary and alternative therapies are being undertaken. Tese include use of vitamin E [8], ginkgo biloba extract [9], and choline alfoscerate [10], which have been tested for treatment of dementia. However, there are no clear indications supporting their efcacy.
Kami Guibi-tang (KGT) is a traditional herbal medicine used in East Asia that is widely used for insomnia and depression [11]. Complementary and alternative medicine (CAM) has been widely used in countries around the world, and herbal medicine is one of the most popular therapy methods in the CAM. Te use of CAM in diferent diseases has been increasing [12], and other clinical trials have shown the efcacy of CAM for diferent diseases [13][14][15][16]. Experimental studies undertaken with KGT have reported that it increases the activity of central nerve cells [17] and improves cognitive impairment by reducing neuronal apoptosis and Aβ accumulation in the hippocampus [18]. It is speculated that the efect of KGT on cognitive function occurs through the reversal of degenerative axonal atrophy and nerve damage caused by the phosphorylation of Aβ and Tau proteins [19][20][21]. An earlier clinical trial reported that KGT improved cognitive function in patients with mild AD [22]. A combination therapy of KGT and AChEI donepezil over a 16-week period was documented to prolong the efect of donepezil [23], and a crossover study reported that it improved cognitive function [24]. A previous pilot study of KGT in patients with a predementia condition known as amnestic mild cognitive impairment (aMCI) found that KGT had signifcant efcacy in improving memory [25].
As described above, KGT had a positive efect on the cognitive function of patients with AD. However, previous studies have limitations in that they were not blinded. Additionally, to date, there has been no study that strictly confrmed the efcacy and safety of KGT through randomized, placebo-controlled, and double-blind studies. Terefore, the aim of this pilot study was to rigorously investigate the efcacy and safety of KGT in improving cognitive function in patients with mild AD.

Study Design.
Tis study was designed as a pilot for a randomized, double-blind, placebo-controlled clinical trial, the protocol for which has been published [26]. Te study was conducted between March 2018 and November 2020 at the Kyung Hee University Hospital at Gangdong, Seoul, Republic of Korea. During a screening period of approximately 2-weeks from the time the prospective participant visited, he/she was evaluated to determine whether they met the criteria for inclusion in the study. All the patients were treated for a period of 24 weeks and followedup thereafter for 4 weeks.
We recruited persons who visited our hospital with complaints of impaired memory. Potential participants voluntarily completing a written informed consent form were screened over a 2-week period according to the inclusion/exclusion criteria (as given below), the Korean Mini-Mental State Examination (K-MMSE), and the Korean Dementia Screening Questionnaire (KDSQ). In the inclusion criteria, the continuous use of AChEI medications was stipulated. Potentially eligible participants completed the Seoul Neuropsychological Screening Battery (SNSB), and those diagnosed with mild AD based on their scores by a neurologist were enrolled in the trial.
Participants were randomly allocated to the treatment or control groups. Te treatment group received KGT granules, and the control group received placebo granules three times a day for 24 weeks. Te dosage of the AChEI medications was maintained in both the groups during the study period. After 24 weeks, the efcacy and safety of the KGT were assessed through comparisons with the baseline. Adverse events were assessed once a week during the medication period and four weeks following the completion of the medication course.
Te changes in the Seoul Neuropsychological Screening Battery-Dementia Version (SNSB-D) total scores and the scores of the fve SNSB-D domains were compared before and after the 24-week treatment/placebo period as the primary outcome. We considered the SNSB subtest scores and scores on the K-MMSE, Short Geriatric Depression Scale (SGDS), Barthel Index of Activities of Daily Living (Barthel ADL), Korean Instrumental Activities of Daily Living (K-IADL), Clinical Dementia Rating (CDR), and Global Deterioration Scale (GDS) as secondary outcomes. In addition, the changes in the Korean version of Quality of Life-Alzheimer's Disease (KQoL-AD) and Caregiver-Administered Neuropsychiatric Inventory (CGA-NPI) scores were compared. We monitored the safety of KGT by regular checks for adverse events and abnormal fndings on vital signs, blood tests, electrocardiogram (ECG), and brain magnetic resonance imaging (MRI).
Te study protocol was approved by the Korean Ministry of Food and Drug Safety (MFDS approval number: 31234) as well as the Institutional Review Board of Kyung Hee University Hospital at Gangdong (IRB approval number: KHNMCOH 2017-11-002-001) and was registered in the Clinical Research Information Service (CRIS approval number: KCT0002904). All participants received a sufcient explanation of the background, method, potential risks and benefts, and provisions for confdentiality before the start of the study following, which they voluntarily flled out the written consent form. All the processes of this clinical study were conducted in compliance with the ethical regulations of the Declaration of Helsinki (South Africa Amendment, 1996) and the Korea Good Clinical Practice (KGCP) guidelines. Tis clinical study complied with the Consolidated Standards of Reporting Trials (CONSORT) statement (Table S2).

Inclusion Criteria.
Participants meeting the following criteria were eligible to participate if they were aged 55-90 years old; complained of impaired memory; were diagnosed with mild AD by a neurologist, based on scores on the SNSB and the criteria of the National Institute of Neurological and Communicative Disorders and Stroke and Alzheimer's Disease and Related Disorders Association (NINCDS-ADRDA), with a CDR of 0.5∼1 point; were constantly taking AChEIs, such as donepezil, rivastigmine, and galantamine, and free of adverse events for the previous four weeks, and cognition-related medications included agents to improve cerebral blood fow and others afecting cognition, such as gliatilin, gliatamin, ginexin, and tanamine; were taking medications such as sleeping pills, antianxiety drugs, antidepressants, antipsychotics, and anticholinergic drugs to stabilize the underlying disease for the previous three months or more, and no indications of probable loss of stability during the 24-week study period; had no problems in communication; no contraindications for MRI.

Exclusion Criteria.
Participants who met any of the following criteria were ineligible to participate; a brain disorder causing neurological symptoms other than cognitive dysfunction; formal diagnosis of Parkinson's disease, Huntington's chorea, Down syndrome, Creutzfeldt-Jakob disease, and other neurodegenerative disorders; cognitive impairment resulting from conditions such as head trauma, hypoxic brain damage, vitamin defciency, brain tumor, encephalitis, neurosyphilis, or mental retardation; cerebrovascular disease documented by MRI; history of convulsive disease other than febrile convulsions in childhood; diagnosed psychiatric disorders such as major depressive disorder, bipolar disorder, schizophrenia, alcoholism, or substance abuse, according to the Diagnostic and Statistical Manual of Mental Disorders, 4 th edition (DSM-IV) criteria; life-threatening physical disabilities requiring immediate treatment; uncontrolled hypertension; heart or renal disease; peripheral edema; gastrointestinal symptoms, such as anorexia, stomach discomfort, nausea, abdominal pain, and diarrhea; use of medications that could induce hypokalemia or myopathy; use of NMDA receptor antagonist, such as memantine; hypersensitivity to the medication used in the study; possibility of pregnancy; clinically signifcant abnormalities in blood chemistry, including levels of serum glutamic pyruvate transaminase (SGPT)/serum glutamic oxaloacetic transaminase (SGOT) being more than twice the normal upper limit, or serum creatinine levels more than 10% above the normal upper limit; participation in any other clinical trial within the previous 4 weeks; illiteracy; considered unsuitable for participation by the investigators.

Termination Criteria.
Participants who met any of the following criteria were stopped in the study; severe adverse events making it impossible to continue the clinical trial; individual considered unsuitable for participation by the investigators; voluntary withdrawal; nonobservance of the protocol, i.e., drug compliance below 80%; decision by the principal investigator.

Recruitment.
Between March 2018 and May 2020, we recruited patients aged 55-90 years with complaints of impaired memory through advertisements on bulletin boards, newspapers, and online media. Potential participants were screened using the K-MMSE and the KDSQ. Potentially eligible participants, who completed the SNSB and those who were diagnosed with mild AD by a neurologist, were enrolled in the study.

Randomization and Blinding.
Participants were assigned to either the treatment group (KGT group) or control group (placebo group) at a ratio of 1 : 1 using a block randomization method with a block size of 4. Te random number table was generated by a researcher who did not participate in the evaluation, and the numbers were assigned sequentially with enrollment. KGT and placebo granules with identical appearance, smell, taste, and properties were produced by the manufacturer (Kyungbang Co., Incheon, Korea). Te placebo consisted of corn starch, lactose, hydroxypropyl cellulose, caramel color (food additives), tartrazine (FD and C Yellow 5), Allura Red AC (FD and C Red 40), and Ssanghwa favor. Te manufacturer assembled the products in the medication kits, each labeled with a study-generated random number. An independent Evidence-Based Complementary and Alternative Medicine pharmacist distributed the kits to the participants in the order of their enrollment. Participants, investigators, pharmacists, and outcome assessors were blinded to the assignment until the end of the trial. Te blinding could be broken according to the approved procedure in case of serious adverse events.

Intervention.
Participants in both the study groups were instructed to ingest the contents of one 3.0 g packet from their medication, KGT, or placebo granules kit (Kyungbang Co., Incheon, Korea) with water three times a day, 30 min after meals, for 24 weeks. Both drugs were in the form of yellow-brown granules. Te components and dosages of KGT are listed in Table 1.

Korean Dementia Screening Questionnaire (KDSQ) and Korean Mini-Mental State Examination (K-MMSE).
Te KDSQ is a cognitive screening test administered to caregivers of patients [27]. It consists of questions related to memory and behavioral disorders, and problems performing normal daily activities that are common in early AD [28].
Te K-MMSE is a simple cognitive function test that evaluates the overall cognitive function and was used to exclude persons with normal cognition, mild cognitive impairment (MCI), or dementia [29]. It is the most widely used cognitive function screening test and is included in the NINCDS-ADRDA diagnostic criteria for AD. Age-and education-specifc standard scores have been developed for normal cognitive function.

Seoul Neuropsychological Screening Battery-Dementia
Version (SNSB-D). Te SNSB was assessed at baseline and at 24 weeks by an independent clinical psychologist to assess the impact of KGT on cognitive function. Te SNSB is a standardized neuropsychological test often used in Korea [30]. Tis study used the revised version, the SNSB-II, which evaluates fve cognitive domains (attention, language ability, memory, spatiotemporal ability, and executive ability) and includes other related tests of cognitive function as subtests [31]. Te details have been included in Table S1 in the Supplementary Material. SNSB-D is a version of the SNSB modifed for patients with AD [32]. Te test results provide a global cognitive function score, which is the sum of fve cognitive domain subtests: attention (17 points, 6%), language ability (27 points, 9%), memory ability (150 points, 50%), spatiotemporal ability (35 points, 12%), and executive ability (70 points, 23%).
Te SGDS is a shortened depression scale designed to evaluate depressive symptoms in the elderly [33]. It contains questions covering complaints of memory loss and cognitive dysfunction, which are often the symptoms of depression in the elderly. Tis scale uses easily understood questions that can be answered with yes or no. Te risk of depression increases with the score; in general, scores of 8 (out of 15) or higher indicate a high risk.
Te Barthel ADL consists of 10 items that evaluate basic daily activities such as feeding, personal toileting, bathing, dressing and undressing, getting on and of the toilet, controlling bladder, controlling bowel, moving from wheelchair to bed and returning, walking on a level surface or propelling a wheelchair if unable to walk, and ascending and descending stairs [34].
Te K-IADL scale evaluates instrumental daily life activities, which are higher than basic daily life activities [35]. It is completed by a caregiver who has witnessed the patient's daily life for at least the previous four weeks and consists of 11 questions on a 0-3 point scale. Topics include shopping, travel (mode of transportation), ability to handle fnances, housekeeping (use of electronic devices), preparing food, ability to use the telephone, responsibility for one's own medication, recent memory, hobbies, watching TV, and fxing around the house.
Te CDR scale evaluates the overall severity of AD by assessing both the cognitive levels and daily living ability [36]. Six domains of memory, orientation, judgment and problem solving, community afairs, home and hobbies, and personal care were evaluated through semistructured interviews with patients and caregivers. Te CDR Global Score (CDR-GS) is based only on the CDR memory score: CDR Te GDS expresses cognitive decline and subsequent functional deterioration on a scale of 1 to 7 [37]. Tis scale describes the degree of cognitive impairment at each stage with specifc examples and diferentiates the initial stages of cognitive impairment in detail. Similar to the CDR, the GDS assesses the severity of dementia and is widely used for clinical research on cognitive drugs and the early diagnosis of dementia.

Korean Version of Quality of Life-Alzheimer's Disease (KQoL-AD).
Te KQoL-AD test assesses the overall quality of life of patients with AD [38]. Its reliability and validity are relatively high in patients with mild AD and can be applied to patients with signifcantly reduced cognitive function. Patients and caregivers separately evaluated physical health, energy level, mood, living situation, memory, relationships with spouse, relatives and friends, self as a whole, chores around the house, leisure activities, fnancial situation, and life as a whole, each on a scale of 1 to 4.

Blood Test, Electrocardiogram, and Brain MRI.
Blood tests were performed at baseline and at 12 and 24 weeks to evaluate safety. Specifcally, blood urea nitrogen, creatinine, SGOT, SGPT, sodium, potassium, chloride, creatine kinase, lactate dehydrogenase, and glucose were monitored to detect abnormalities in liver function, renal function, electrolytes, and heart enzyme levels. Apolipoprotein E (ApoE) genotyping was performed to test for the ε4 allele, which carries an increased risk for AD [40]. An ECG was performed at baseline, 12 weeks, and 24 weeks to monitor drug safety. Brain MRI was performed at baseline and after 24 weeks. We checked whether there were any structural abnormalities in the brain through 3D T1 weighted images, 2D T2 weighted images, and fuidattenuated inversion recovery (FLAIR) images.

Sample Size Calculation.
Previous clinical studies reporting on the efcacy of KGT in improving cognitive function in patients with mild AD did not use methodology equivalent to that of the present study in terms of study design and disease state. Tus, there were no available data for calculating the sample size. Since this was a pilot study, the number of study participants was based on the number of patients that could be recruited and the projected cost of the overall study. In general, 20-40 people per group were suggested as the appropriate number of participants in a pilot study [41]. Accordingly, the target number for this study was set to 30 per group, with the goal of 38 enrolled participants per group to accommodate a dropout rate of up to 20%.

Statistical Analysis.
All data were entered into Microsoft Excel 2016 (Microsoft Co., Redmond, USA) and analyzed using SAS statistical software (version 9.4; SAS Institute, Cary, NC, USA). Te analysis was performed as intention-to-treat using those who took the test drug at least once after randomization.
Demographic data and cognitive function assessment results, such as the SNSB scores, were tested using the chisquared test or Fisher's exact test for diferences between groups at baseline and to compare prognostic variables.
Diferences in representative values were tested by the Wilcoxon rank-sum test. For within-group comparisons of continuous variables before and after the study period, paired tests were performed using the Wilcoxon signed-rank test. Statistical analyses were performed at a signifcance level of 5%, so a p value less than 0.05 was considered statistically signifcant. All descriptive statistics of the results are expressed as mean ± standard deviation (SD) or median (IQR) for continuous variables and frequency (%) for categorical variables.

Participants.
A total of 154 individuals expressed their intention to participate in the study. Prospective participants completed telephone screening and the K-MMSE and KDSQ tests. Te SNSB was subsequently administered to 20 potentially eligible individuals whose screening results indicated possible mild AD and who provided written informed consent. Of these 20 patients, two did not meet the inclusion criteria and two withdrew their consent. Based on the SNSB results, 16 participants were diagnosed with mild AD by a neurologist and were enrolled in the study. Among them, nine were randomized to the treatment group and seven to the control group. Two participants in the treatment group did not complete the study; one withdrew and the other had to be excluded from the study due to violation of the study protocol. Both were included in the intention-totreat analysis. Seven participants each in the treatment and the control group completed the trial ( Figure 1).

General and Clinical Features.
Tere were no signifcant diferences between the KGT and control participants in terms of the general characteristics such as age, sex, education level, and presence of the ApoE ε4 allele at baseline. Te results of the SNSB-D and other cognitive tests were not signifcantly diferent between the two groups at baseline.

Evidence-Based Complementary and Alternative Medicine
However, there was a signifcant diference in mean SGDS scores, with scores of 6.29 ± 3.68 and 1.83 ± 2.64 for the KGT and control groups, respectively (p � 0.04, Table 2).

Efcacy Outcome
While the mean score for the SNSB-D in the KGT group showed a nonsignifcant decrease from 119.86 ± 30.38 at baseline to 112.93 ± 31.58 at 24 weeks (p � 0.69), the score in the control group showed a nonsignifcant increase from 93.79 ± 23.58 at baseline to 96.57 ± 19.44 at 24 weeks (p � 0.63). Te changes in the mean scores were not signifcantly diferent between the two groups (p � 0.71). Analysis of the changes in the scores in the fve domains of the SNSB-D showed no signifcant diferences either within or between the groups (Table 3).  (Table 4).

KQoL-AD.
Scores on the KQoL-AD did not show a signifcant change in the KGT group, either when the patient self-evaluated (31.14 ± 5.58 to 28.57 ± 4.86, p � 0. 16) or when the caregiver provided the evaluation (27.86 ± 5.11 to 27.00 ± 7.48, p � 0.58). Changes between baseline and 24 weeks in the control group were not signifcant when the patient self-evaluated (34.29 ± 6.60 to 34.14 ± 5.21, p � 1.00) or when the caregiver provided the evaluation (30.57 ± 7.52, 29.86 ± 4.56, p � 1.00). Tere were no signifcant diferences between the treatment and control groups for either the caregiver or patient evaluations (p � 0.61 and p � 0.66, respectively, Table 4).

Comparative Analysis with aMCI Group.
A pilot study was conducted on aMCI with the same basic protocol as in the present study [25]. With the consent of the author, we compared the data from that study on patients with aMCI receiving KGT (n � 16) for 24 weeks with the present datapatients with mild AD receiving KGT (n � 7). Te SNSB-D total score and memory domain score were signifcantly higher in the aMCI group than in the mild AD group (p � 0.04 and 0.02, respectively). In addition, the Rey recall score in the memory domain, contrasting program score, Luria loop score, and word fuency (animal naming) scores in the frontal and executive function domains showed a signifcant diference (p � 0.03, 0.03, 0.05, and 0.04, respectively). Tere was no signifcant diference between the two groups in the scores of the other subtests (Table 5). Te amount of change was signifcantly diferent between the aMCI and mild AD groups for the K-MMSE and K-IADL scores (p � 0.01 and 0.02, respectively). No signifcant diferences between the two groups with respect to the SGDS, Barthel-ADL, GDS, and CDR scores were observed (Table 6).

Discussion
To our knowledge, this is the frst randomized, double-blind, placebo-controlled study to explore the efcacy of KGT in improving the cognitive function and safety of patients with mild AD. We expanded the methods used in previous studies of KGT to include SNSB, KQoL-AD, and CGA-NPI. During the 24-week treatment period, changes in the SNSB-D and other indices were not signifcantly diferent between the treatment and placebo groups. Te SNSB covers a wide range of difculty levels, allowing an in-depth evaluation of each cognitive domain. However, the test takes about 2 h, which can be a burden for elderly people. Participants experiencing severe cognitive decline, are elderly, or who are uncooperative will have more difculty completing the test [42]. In this study, the SNSB-D score tended to decrease over time in the KGT group. Tis is likely a refection of several participants in the KGT group with    Evidence-Based Complementary and Alternative Medicine  Evidence-Based Complementary and Alternative Medicine high initial scores for negative items in the CGA-NPI, such as apathy/indiference and depression/discouragement. Te mean K-MMSE scores decreased signifcantly over the study period in both groups, although the comparison between them was not signifcant. Most studies on mild AD have used MMSE-1 as an evaluation tool [22][23][24]. On an average, the MMSE-1 scores in patients with mild AD decrease by 3.8 points per year [43]. In previous KGT treatment studies, the MMSE-1 score was maintained or increased by an average of 1 point. In contrast, it has been reported that among patients with mild AD, higher initial MMSE-1 scores are associated with lower drug treatment results [44]. Te initial MMSE-1 scores of this study averaged 21-22 points, higher than those reported for clinical studies of KGT [22,24], and this diference in cognitive levels as measured by the MMSE-1 may have infuenced the treatment efcacy.
Additionally, we compared the efects of KGT on the cognitive function of patients with aMCI and mild AD and analyzed the diferences between the two groups. According to a previous preliminary clinical study [25], the KGT treatment group with aMCI showed a signifcant improvement in the SNSB-D score. In this study, compared to the mild AD group, KGT was associated with a signifcant improvement in the SNSB-D score in the MCI stage than in the mild AD stage, particularly in the memory domain of SNSB-D. Te results showed that KGT signifcantly improved the Rey recall score in the memory domain, contrasting program, the Luria loop, and the word fuency score in the frontal and executive function domains showed signifcant diferences.
Te Rey recall test is a memory recall test belonging to the Rey-Osterrieth Complex Figure Test, a representative test tool for spatiotemporal cognitive function, and checks whether the stimulus is accurately input by the patient as they must draw the same as the given picture stimulus, by remembering that was viewed. It is also a useful tool for evaluating the executive function related to frontal lobe function by redrawing it. In patients with mild AD, the Rey recall score of the SNSB is related to brain gray matter volume, which is known to be associated with impaired memory of language and visual information due to atrophy of the superior temporal gyrus [45]. Te contrasting program test evaluates the ability to control movements and learn rules. If the tester raised two fngers, the subject was instructed to raise one fnger, and if the tester raised one fnger, the subject was instructed to raise two fngers. If frontal lobe dysfunction is severe, the subject follows the examiner's fnger. In this case, if response inhibition is not performed, it is judged that there is defective response inhibition [46]. Te Luria loop test is used to determine if there is internal perseveration by drawing several loops with three rings [47]. Te word fuency test, the animal naming, evaluates semantic word fuency by making the animal's name words spoken as much as possible within 1 min. Semantic fuency is related to semantic memory, which is applied to words and meanings, and to components of longterm memory. Te impairment of the semantic memory test means there is a lack of access to item knowledge or loss of representational knowledge [48]. In general, it is known that frontal lobe function is maintained in the early stages of AD, but studies have shown that frontal executive function abnormalities appear at an early stage [49]. Executive function is concerned with the control and regulation of cognitive processes and goal-directed, future-oriented behavior [50].
Te signifcant improvement in the memory and frontal function test scores in the aMCI group showed that KGT was more efective in the MCI stage than in the mild AD stage. It is thought that KGT has the potential to have a positive efect on neuropathology that can progress to AD, such as temporal lobe atrophy, as well as improving memory, language, and visual information processing ability. However, we found that KGT was efective in improving cognitive function in patients with aMCI but not in those with mild AD. Terefore, it is necessary to consider the reasons for these diferences. First, participants in the aMCI pilot study took KGT alone, whereas our study participants took KGT with AChEI. Tese conventional drugs may have masked the efects of KGT, but discontinuing an AChEI to participate in a study creates an ethical problem. Second, because aMCI is a predementia stage before neurodegenerative changes have occurred, the efcacy of KGT may have been better expressed at this stage than at the stage of mild AD, in which the degenerative changes have already begun, and recovery cannot be expected. Finally, approximately 20% of patients with aMCI naturally improve to normal [51]. It is remarkable that KGT showed signifcant efects on cognitive function improvement in patients with aMCI before the treatment guidelines for KGT in MCI have been established. Terefore, these results indicate the need for initiating KGT treatment at earlier stages of cognitive impairment.
Te efects of KGT have been demonstrated in experimental studies in mice. In a 2008 study on an Aβ-induced mouse model of AD, ingestion of Guibi-tang for 3 days improved memory acquisition, memory retention, and object recognition [19]. In a 2011 5FXAD mouse study, KGT improved object recognition and reduced the number of amyloid plaques in the frontal cortex and hippocampus [20]. In another study, KGT ameliorated Aβ-induced tau phosphorylation and axonal atrophy [21]. Previous clinical studies testing the efects of KGT on cognitive function include clinical trials and retrospective chart reviews. All studies used the MMSE-1 as an evaluation tool. In a clinical trial reported by Higashi et al., the MMSE-1 score was signifcantly higher than that in the nontreatment group and the group receiving Goshajinkigan [22]. A crossover test by Watari et al., reported that MMSE-1 scores were signifcantly increased after 16 weeks of combination therapy with donepezil compared to 16 weeks of donepezil alone [23]. In a retrospective study, the MMSE-1 scores were lower in patients receiving donepezil alone than in those receiving donepezil plus KGT [24].
KGT is derived from 15 herbs and plants. Studies have documented positive efects on the cognitive function of each component. Polygalae radix, the main ingredient of KGT, acts on the cholinergic system of the CNS66, and exerts efects on neurodegenerative pathologies through an increase in acetylcholine levels, such as improving cognitive function, neuroprotection against Aβ, and suppression of neuroinfammation [52]. Gardeniae fructus, another key ingredient, promotes cholinergic neurotransmission and improves memory [53]. Moutan cortex inhibits the aggregation of Aβ proteins [54]. Poria sclerotium is a long-term synergistic efector of the hippocampus [55] and inhibits acetylcholinesterase [56]. Zizyphi fructus reduced the number of activated microglia and astrocytes observed after Aβ injection [57]. Tis combination of properties clarifes the possible mechanisms of KGT efcacy.
Tis study has several limitations. First, we were able to recruit only fewer participants than planned. Tis is partly due to the inclusion criteria, which were stricter than those in previous studies. In addition, the latter part of the study period overlapped with the COVID-19 pandemic, making it more difcult to recruit participants. Te resulting small sample size led to low statistical power, afecting our ability to detect signifcant diferences. Several of the limitations mentioned below refect the small sample size. Second, despite the use of randomization to minimize disturbance bias, an imbalance in the SGDS score occurred at baseline. Specifcally, participants taking antidepressants were assigned to the KGT group. It is known that 30%-50% of patients with AD sufer from depression [58], but depression is not the only risk factor for cognitive decline. Te relationship between depression and AD is complex; the conditions may appear independently, or depression may appear as a secondary reaction to cognitive impairment. Since antidepressant medications can afect cognitive function, the imbalance in assignment may have afected the results. Tree participants with SGDS scores of 8 or higher were assigned to the KGT group, and two of them improved to the normal range after treatment. Tird, AD was only diagnosed by SNSB at the screening stage, rather than by imaging tests such as amyloid PET, tau PET, or neurodegeneration MRI. Fourth, the SNSB-D score tended to decrease in the KGT group. Tere were several participants with initial high apathy/indiference and agitation/aggression scores on the CGA-NPI. Tese participants were uncooperative and complained of difculty completing longer tests such as the SNSB. Fifth, this study examined KGT as an add-on therapy for AChEI and could not assess its efects when used alone. Te previous aMCI pilot study used KGT alone and found efcacy in improving cognitive function. Finally, the study was not designed with a follow-up observation period, so any long-term efects of the treatment on disease progression could not be determined.
Tis study was a productive frst attempt to observe the efcacy and safety of KGT in mild AD, which remains incurable. Te results of this study can be used as reference data for future large-scale clinical trials on the efcacy of KGT for treating cognitive diseases.

. Conclusions
KGT treatment over 24 weeks did not have a signifcant impact on overall cognitive function as measured by the SNSB in patients with mild AD. KGT was associated with signifcant improvements in the memory domain in the aMCI group compared to the mild AD group, and no adverse events were reported. Te results of this pilot study will be valuable in designing future larger-scale clinical trials on the efcacy of KGT for treating cognitive diseases.

Data Availability
Te datasets used to support the fndings of this study are available from the corresponding author upon request.

Conflicts of Interest
Te authors declare that there are no conficts of interest.